Approximately one-third of the world's population is infected with Mycobacterium tuberculosis, the causative agent of tuberculosis. While the majority of these infections do not progress immediately to active disease, tuberculosis results in about 3 million deaths each year. In the United States, there has been a significant increase in the incidence of tuberculosis in recent years, in large part due to an increase in HIV infection. This increase is accompanied by the alarming appearance of drug resistant and multiple drug resistant strains of tuberculosis. There is clearly a need for new antituberculosis drugs, better and more effective vaccines and rapid methods for diagnosis and drug susceptibility determination. Effective control of tuberculosis requires an understanding of the genetic basis of M. tuberculosis virulence, mechanisms of gene expression and its regulation, and specific tools for the genetic manipulation of the mycobacteria. However, it is only very recently that a genetic system for mycobacteria has been established and these questions have yet to be fully addressed. Viruses have proven to be valuable aids in the exploration of a variety of organismal genetic systems. It is expected that a study of mycobacterial viruses will both enhance our understanding of mycobacterial genetics and provide tools for genetic manipulation of pathogenic mycobacteria. Previous studies have established mycobacteriophage L5 as the best characterized of the mycobacterial viruses and validated the utility of this approach. This project proposes to elucidate the mechanisms of gene expression and regulation in L5 and its mycobacterial hosts; this information will be used for the construction of novel diagnostic luciferase reporter phages and to provide tools for the construction of recombinant antituberculosis vaccines. These studies will also contribute to the repertoire of tools available for genetic manipulation of the mycobacteria.